Although primary transplantation has evolved into an effective therapy for end-stage lung disease, the balance between organ supply and demand greatly limits its applicability. Pulmonary xenotransplantation using swine lungs in humans could provide a solution to this problem. However, pulmonary xenografts are rapidly reflected by mechanisms distinct from those causing hyperacute rejection of the heart or kidney. The long-term objective of this project is to determine the mechanisms responsible for pulmonary xenograft injury, develop strategies which will prevent this injury and to enable the clinical application of pulmonary xenotransplantation. Although the mechanisms responsible for rejection of the cardiac xenograft are dependent upon the binding of xenoreactive antibody to the Gal(1-3)Ga1 epitope with subsequent complement activation, the importance of this interaction in lung xenografts is unclear. The aims of this proposal are as follows: (1) define the role of the pulmonary intravascular macrophage and other resident macrophage and other resident macrophages within the lung in inducing pulmonary vasoconstriction, endothelial cell activation and initiation of coagulation; (2) to define the role of classical complement pathway activation, particularly anaphylatoxins and xenoreactive antibody in pulmonary xenograft injury; and (3) characterize the role of tissue factor initiated coagulation and inadequate regulation of coagulation in pulmonary xenograft injury particularly with regards to endothelial cell activation and augmentation of inflammation. The experiments outlined in this proposal will (1) deplete resident macrophages, both the intravascular and alveolar macrophages; (2) use of anti-C5a monoclonal antibodies and other inhibitors of the complement system; (3) depletion of Gal antibody and to use lungs of the Gal epitope; and (4) blockers of tissue factor dependent coagulation, swine vWF platelet interaction and regulator of the coagulation system that are effective in a swine to primate model. It is anticipated that these studies will further delineate the mechanisms responsible for the early pulmonary xenograft injury and to begin to allow for examination of the mechanisms responsible for acute vascular rejection. Strategies developed from these studies should form the basis for the application of pulmonary xenotransplantation to the treatment of patients with pulmonary failure.